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1.
World J Microbiol Biotechnol ; 35(9): 132, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31432260

RESUMO

This paper aims to characterize halophilic bacteria inhabiting Algerian Saline Ecosystems (Sebkha and Chott) located in arid and semi-arid ecoclimate zones (Northeastern Algeria). In addition, screening of enzymatic activities, heavy metal tolerance and antagonistic potential against phytopathogenic fungi were tested. A total of 74 bacterial isolates were screened and phylogenetically characterized using 16S rRNA gene sequencing. The results showed a heterogeneous group of microorganisms falling within two major phyla, 52 strains belonging to Firmicutes (70.2%) and 22 strains (30.8%) of γ-Proteobacteria. In terms of main genera present, the isolates were belonging to Bacillus, Halobacillus, Lentibacillus, Oceanobacillus, Paraliobacillus, Planomicrobium, Salicola, Terribacillus, Thalassobacillus, Salibacterium, Salinicoccus, Virgibacillus, Halomonas, Halovibrio, and Idiomarina. Most of the enzymes producers were related to Bacillus, Halobacillus, and Virgibacillus genera and mainly active at 10% of growing salt concentrations. Furthermore, amylase, esterase, gelatinase, and nuclease activities ranked in the first place within the common hydrolytic enzymes. Overall, the isolates showed high minimal inhibitory concentration values (MIC) for Ni2+ and Cu2+ (0.625 to 5 mM) compared to Cd2+ (0.1 to 2 mM) and Zn2+ (0.156 to 2 mM). Moreover, ten isolated strains belonging to Bacillus, Virgibacillus and Halomonas genera, displayed high activity against the pathogenic fungi (Botrytis cinerea, Fusarium oxyporum, F. verticillioides and Phytophthora capsici). This study on halophilic bacteria of unexplored saline niches provides potential sources of biocatalysts and novel bioactive metabolites as well as promising candidates of biocontrol agents and eco-friendly tools for heavy metal bioremediation.


Assuntos
Antibiose , Bactérias/isolamento & purificação , Bactérias/metabolismo , Biota , Microbiologia Ambiental , Salinidade , Argélia , Bactérias/classificação , Bactérias/genética , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Fungos/crescimento & desenvolvimento , Hidrolases/análise , Metais Pesados/metabolismo , Metais Pesados/toxicidade , Testes de Sensibilidade Microbiana , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
2.
Pol J Microbiol ; 68(2): 193-201, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31250589

RESUMO

The present study aimed to isolate the high-efficiency petrol metabolizing thermophilic bacteria from petrol contaminated soil samples. Isolation was carried out through enrichment culture, serial dilution and pour plate methods using the petrol supplemented minimal salt media. The isolated bacteria were analyzed to document growth behavior, petrol removal efficiencies, antibiotic resistance profile, and biochemical characteristics. The 16S rRNA based phylogenetic analysis helped to reveal the identity of isolated bacterial species and construct the phylogenetic trees. Total nine bacteria were isolated, out of which three (IUBP2, IUBP3, IUBP5) were identified as Brevibacillus formosus, one (IUBP1) was found similar to Brevibacillus agri, four (IUBP7, IUBP8, IUBP13, and IUBP14) shared homology with Burkholderia lata, and one (IUBP15) with Burkholderia pyrrocinia. All the isolates were fast growing and exhibited considerable petrol degradation potential. The highest petrol removal efficiency (69.5% ± 13.44/6 days) was recorded for the strain IUBP15 at a petrol concentration of 0.1% (v/v). All bacteria studied (100%) were positive for esculinase and phosphatase. Many strains exhibited positive responses for arginine dehydrolase (22%), ß-naphthylamidase (11%), ß-D-glucosaminide (33%), mannitol (55%), sorbitol (66%) and inulin (88%) fermentation test. While all were sensitive to the antibiotics, some of them were found resistant against chloramphenicol and oxacillin. The remarkable biochemical characteristics and considerable petrol removal potential (40-70%) highlights utilization of the bacteria isolated for petrol bioremediation, mineralization of organophosphates, dairy and food industry, and also as biofertilizers and biocontrol agents.The present study aimed to isolate the high-efficiency petrol metabolizing thermophilic bacteria from petrol contaminated soil samples. Isolation was carried out through enrichment culture, serial dilution and pour plate methods using the petrol supplemented minimal salt media. The isolated bacteria were analyzed to document growth behavior, petrol removal efficiencies, antibiotic resistance profile, and biochemical characteristics. The 16S rRNA based phylogenetic analysis helped to reveal the identity of isolated bacterial species and construct the phylogenetic trees. Total nine bacteria were isolated, out of which three (IUBP2, IUBP3, IUBP5) were identified as Brevibacillus formosus, one (IUBP1) was found similar to Brevibacillus agri, four (IUBP7, IUBP8, IUBP13, and IUBP14) shared homology with Burkholderia lata, and one (IUBP15) with Burkholderia pyrrocinia. All the isolates were fast growing and exhibited considerable petrol degradation potential. The highest petrol removal efficiency (69.5% ± 13.44/6 days) was recorded for the strain IUBP15 at a petrol concentration of 0.1% (v/v). All bacteria studied (100%) were positive for esculinase and phosphatase. Many strains exhibited positive responses for arginine dehydrolase (22%), ß-naphthylamidase (11%), ß-D-glucosaminide (33%), mannitol (55%), sorbitol (66%) and inulin (88%) fermentation test. While all were sensitive to the antibiotics, some of them were found resistant against chloramphenicol and oxacillin. The remarkable biochemical characteristics and considerable petrol removal potential (40­70%) highlights utilization of the bacteria isolated for petrol bioremediation, mineralization of organophosphates, dairy and food industry, and also as biofertilizers and biocontrol agents.


Assuntos
Bactérias/isolamento & purificação , Bactérias/metabolismo , Petróleo/metabolismo , Microbiologia do Solo , Poluentes do Solo/metabolismo , Antibacterianos/farmacologia , Bactérias/classificação , Bactérias/genética , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Farmacorresistência Bacteriana , Enzimas/análise , Testes de Sensibilidade Microbiana , Poluição por Petróleo , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
3.
Pol J Microbiol ; 68(2): 203-209, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31250590

RESUMO

The composition of vagina lactic acid bacteria (LAB) differs within the different ethnic group. This study is aimed at determining the prevalence of LAB with their antimicrobial properties in Nigerian women's vagina during different stages of the menstrual cycle. Microorganisms were isolated from vaginal swabs of ten Nigerian women during different stages of the menstrual cycle and identified by partial sequencing of the 16S rRNA gene. The antimicrobial properties of the LAB were tested against the multidrug-resistant uropathogens. The prevalence of LAB was higher during ovulation period while during menstruation period, it declined. Twenty-five LAB isolates were identified as three species, namely: Lactobacillus plantarum (15), Lactobacillus fermentum (9), Lactobacillus brevis (1) and one acetic acid bacteria - Acetobacter pasteurianus. The LAB had antimicrobial activities against the three uropathogens with zones of inhibition from 8 to 22 mm. The presence of LAB inhibits the growth of Staphylococcus sp. GF01 also in the co-culture. High LAB counts were found during ovulation period with L. plantarum as a dominant species while during menstruation, there was a decrease in the LAB counts. The isolated LAB has antimicrobial properties against the urogenital pathogens tested thus exhibiting their potential protective role against uropathogens.The composition of vagina lactic acid bacteria (LAB) differs within the different ethnic group. This study is aimed at determining the prevalence of LAB with their antimicrobial properties in Nigerian women's vagina during different stages of the menstrual cycle. Microorganisms were isolated from vaginal swabs of ten Nigerian women during different stages of the menstrual cycle and identified by partial sequencing of the 16S rRNA gene. The antimicrobial properties of the LAB were tested against the multidrug-resistant uropathogens. The prevalence of LAB was higher during ovulation period while during menstruation period, it declined. Twenty-five LAB isolates were identified as three species, namely: Lactobacillus plantarum (15), Lactobacillus fermentum (9), Lactobacillus brevis (1) and one acetic acid bacteria ­ Acetobacter pasteurianus. The LAB had antimicrobial activities against the three uropathogens with zones of inhibition from 8 to 22 mm. The presence of LAB inhibits the growth of Staphylococcus sp. GF01 also in the co-culture. High LAB counts were found during ovulation period with L. plantarum as a dominant species while during menstruation, there was a decrease in the LAB counts. The isolated LAB has antimicrobial properties against the urogenital pathogens tested thus exhibiting their potential protective role against uropathogens.


Assuntos
Antibiose , Lactobacillales/isolamento & purificação , Lactobacillales/fisiologia , Ciclo Menstrual , Vagina/microbiologia , Acetobacter/classificação , Acetobacter/genética , Acetobacter/isolamento & purificação , Acetobacter/fisiologia , Adulto , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Feminino , Humanos , Lactobacillales/classificação , Lactobacillales/genética , Nigéria , Filogenia , Prevalência , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Staphylococcus/crescimento & desenvolvimento , Adulto Jovem
4.
Pol J Microbiol ; 68(2): 247-254, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31250595

RESUMO

Gastric microbiota provides a biological barrier against the invasion of foreign pathogens from the oral cavity, playing a vital role in maintaining gastrointestinal health. Klebsiella spp. of oral origin causes various infections not only in gastrointestinal tract but also in other organs, with Klebsiella pneumoniae serotype K1 resulting in a liver abscess (KLA) through oral inoculation in mice. However, the relationship between gastric microbiota and the extra-gastrointestinal KLA infection is not clear. In our study, a 454 pyrosequencing analysis of the bacterial 16S rRNA gene shows that the composition of gastric mucosal microbiota in mice with or without KLA infection varies greatly after oral inoculation with K. pneumoniae serotype K1 isolate. Interestingly, only several bacteria taxa show a significant change in gastric mucosal microbiota of KLA mice, including the decreased abundance of Bacteroides, Alisptipes and increased abundance of Streptococcus. It is worth noting that the abundance of Klebsiella exhibits an obvious increase in KLA mice, which might be closely related to KLA infection. At the same time, the endogenous antibiotics, defensins, involved in the regulation of the bacterial microbiota also show an increase in stomach and intestine. All these findings indicate that liver abscess caused by K. pneumoniae oral inoculation has a close relationship with gastric microbiota, which might provide important information for future clinical treatment.Gastric microbiota provides a biological barrier against the invasion of foreign pathogens from the oral cavity, playing a vital role in maintaining gastrointestinal health. Klebsiella spp. of oral origin causes various infections not only in gastrointestinal tract but also in other organs, with Klebsiella pneumoniae serotype K1 resulting in a liver abscess (KLA) through oral inoculation in mice. However, the relationship between gastric microbiota and the extra-gastrointestinal KLA infection is not clear. In our study, a 454 pyrosequencing analysis of the bacterial 16S rRNA gene shows that the composition of gastric mucosal microbiota in mice with or without KLA infection varies greatly after oral inoculation with K. pneumoniae serotype K1 isolate. Interestingly, only several bacteria taxa show a significant change in gastric mucosal microbiota of KLA mice, including the decreased abundance of Bacteroides, Alisptipes and increased abundance of Streptococcus. It is worth noting that the abundance of Klebsiella exhibits an obvious increase in KLA mice, which might be closely related to KLA infection. At the same time, the endogenous antibiotics, defensins, involved in the regulation of the bacterial microbiota also show an increase in stomach and intestine. All these findings indicate that liver abscess caused by K. pneumoniae oral inoculation has a close relationship with gastric microbiota, which might provide important information for future clinical treatment.


Assuntos
Biota , Disbiose/complicações , Mucosa Gástrica/microbiologia , Infecções por Klebsiella/complicações , Abscesso Hepático/complicações , Animais , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Modelos Animais de Doenças , Infecções por Klebsiella/microbiologia , Infecções por Klebsiella/patologia , Abscesso Hepático/microbiologia , Abscesso Hepático/patologia , Camundongos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
5.
Food Chem ; 292: 336-345, 2019 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-31054683

RESUMO

Biochemical effects of the water soluble fiber fraction of sugarcane bagasse (BSF) fermented in the colon was examined to evaluate its potential health promoting effects. A feeding experiment involving Fischer 344 rats, was conducted with 3 experimental diets containing, cellulose (CON), a commercial xylo-oligosaccharide (XYO) and BSF (BGS). Cumulative feed intake was significantly lower in XYO group while cecal weight was significantly higher. Acetic and propionic acid contents in the cecal content were significantly higher in the BGS and XYO, respectively. Total short chain fatty acid content was significantly higher in BGS and XYO resulting significantly lower cecal pH. Beneficial bacteria such as Bifidobacterium, Blautia, Akkermansia and Roseburia abundance was significantly higher in the XYO and BGS groups. Further, mucin and immunoglobulin-A contents were significantly higher in BGS group compared to CON group. Thus, BSF exhibited its ability to enhance the intestinal and systemic health upon fermentation in the colon.


Assuntos
Bactérias/metabolismo , Ceco/microbiologia , Fibras na Dieta/análise , Saccharum/metabolismo , Amônia/análise , Animais , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Bifidobacterium/genética , Bifidobacterium/crescimento & desenvolvimento , Ceco/química , Ceco/metabolismo , Celulose/química , Cromatografia Líquida de Alta Pressão , DNA Bacteriano/química , DNA Bacteriano/metabolismo , Ácidos Graxos Voláteis/análise , Ácidos Graxos Voláteis/química , Masculino , Modelos Animais , Oligossacarídeos/química , Análise de Componente Principal , Ratos , Ratos Endogâmicos F344 , Água/química
6.
Nat Commun ; 10(1): 2194, 2019 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-31097704

RESUMO

Although the physical properties of chromosomes, including their morphology, mechanics, and dynamics are crucial for their biological function, many basic questions remain unresolved. Here we directly image the circular chromosome in live E. coli with a broadened cell shape. We find that it exhibits a torus topology with, on average, a lower-density origin of replication and an ultrathin flexible string of DNA at the terminus of replication. At the single-cell level, the torus is strikingly heterogeneous, with blob-like Mbp-size domains that undergo major dynamic rearrangements, splitting and merging at a minute timescale. Our data show a domain organization underlying the chromosome structure of E. coli, where MatP proteins induce site-specific persistent domain boundaries at Ori/Ter, while transcription regulators HU and Fis induce weaker transient domain boundaries throughout the genome. These findings provide an architectural basis for the understanding of the dynamic spatial organization of bacterial genomes in live cells.


Assuntos
Cromossomos Bacterianos/química , DNA Bacteriano/química , DNA Circular/química , Escherichia coli/genética , Genoma Bacteriano , Proteínas Cromossômicas não Histona/metabolismo , Cromossomos Bacterianos/metabolismo , Replicação do DNA , DNA Bacteriano/metabolismo , DNA Circular/metabolismo , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Microscopia Intravital/instrumentação , Microscopia Intravital/métodos , Microscopia de Fluorescência/instrumentação , Microscopia de Fluorescência/métodos , Conformação de Ácido Nucleico , Análise de Célula Única/instrumentação , Análise de Célula Única/métodos
7.
Molecules ; 24(9)2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-31052562

RESUMO

The role of local DNA structures in the regulation of basic cellular processes is an emerging field of research. Amongst local non-B DNA structures, the significance of G-quadruplexes was demonstrated in the last decade, and their presence and functional relevance has been demonstrated in many genomes, including humans. In this study, we analyzed the presence and locations of G-quadruplex-forming sequences by G4Hunter in all complete bacterial genomes available in the NCBI database. G-quadruplex-forming sequences were identified in all species, however the frequency differed significantly across evolutionary groups. The highest frequency of G-quadruplex forming sequences was detected in the subgroup Deinococcus-Thermus, and the lowest frequency in Thermotogae. G-quadruplex forming sequences are non-randomly distributed and are favored in various evolutionary groups. G-quadruplex-forming sequences are enriched in ncRNA segments followed by mRNAs. Analyses of surrounding sequences showed G-quadruplex-forming sequences around tRNA and regulatory sequences. These data point to the unique and non-random localization of G-quadruplex-forming sequences in bacterial genomes.


Assuntos
Bactérias/genética , DNA Bacteriano/química , Quadruplex G , Genoma Bacteriano , Humanos , Conformação de Ácido Nucleico , Filogenia
8.
Korean J Parasitol ; 57(2): 167-173, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31104409

RESUMO

Rickettsial infections (Rickettsioses) are the causes of acute fever found in Thailand. It is classified as acute febrile illnesses transmitted by bloodsucking arthropod vectors (tick, flea, and chigger). This research investigated pathogens of scrub typhus in vectors from Bangkaew District, Phatthalung Province. A total of 303 pools of vector samples were ticks (Rhipicephalus sanguineus, R. microplus, and Haemaphysalis sp.), fleas (Ctenocephalides felis orientis, C. f. felis, and C. canis), and chiggers (Leptotrombidium deliense, Aschoschoengastia indica, Blankaartia acuscutellaris and Walchia disparunguis pingue) collected from reservoir hosts (dogs and rodents). The 17 and 56 kDa gene of Rickettsia causing scrub typhus were found in 29% of ticks and 98% of flea. DNA sequence analysis reveeled the detected strains were R. asembonensis and Rickettsia sp. cf1 and 5.The chiggers, 1%, were infected with Rickettsia strain TA763, a pathogen of scrub typhus.


Assuntos
Vetores Artrópodes/microbiologia , Orientia tsutsugamushi/isolamento & purificação , Rickettsia/isolamento & purificação , Sifonápteros/microbiologia , Carrapatos/microbiologia , Trombiculidae/microbiologia , Animais , DNA Bacteriano/química , DNA Bacteriano/genética , Cães/parasitologia , Orientia tsutsugamushi/genética , Rickettsia/classificação , Rickettsia/genética , Roedores/parasitologia , Análise de Sequência de DNA , Tailândia
9.
Exp Parasitol ; 201: 21-25, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31029698

RESUMO

Anaplasma phagocytophilum, the bacterial pathogen responsible for tick-borne fever and human granulocytic anaplasmosis, can seriously affect the health of humans and a wide range of other mammals. In this study, we developed a recombinase polymerase amplification (RPA) assay to detect A. phagocytophilum in clinical samples. Following alignment of the relevant DNA sequences, a pair of specific primers based on the 16S rRNA gene was designed to specifically detect A. phagocytophilum. The assay was performed at a constant temperature of 38 °C for 30 min, with a final primer concentration of 0.4 µM. The specificity of the primers was confirmed when DNA from A. phagocytophilum was used as the positive control, and DNA from other related pathogens were used as the negative controls, with ddH2O acting as the blank control. The results showed that the primers did not cross-react with DNA from the other related pathogens. The assay's detection limit was 1.77 × 10-5 ng/µl, a 10 × higher sensitivity level than that determined for nested PCR. The RPA assay's performance was evaluated using 44 clinical samples, and the prevalence results for A. phagocytophilum were found to not differ significantly between the RPA assay and the nested PCR. Thus, we have developed a specific, sensitive, rapid and cost-effective RPA method, requiring only a water bath, for the detection of A. phagocytophilum. The assay should be especially useful in resource-limited areas where access to laboratory equipment is limited.


Assuntos
Anaplasma phagocytophilum/isolamento & purificação , Ehrlichiose/veterinária , Reação em Cadeia da Polimerase/veterinária , Doenças dos Ovinos/microbiologia , Anaplasma phagocytophilum/genética , Animais , Análise Custo-Benefício , DNA Bacteriano/sangue , DNA Bacteriano/química , DNA Bacteriano/isolamento & purificação , DNA Polimerase Dirigida por DNA , Ehrlichiose/diagnóstico , Ehrlichiose/microbiologia , Sistemas Automatizados de Assistência Junto ao Leito/economia , Sistemas Automatizados de Assistência Junto ao Leito/normas , Reação em Cadeia da Polimerase/economia , Reação em Cadeia da Polimerase/normas , RNA Ribossômico 16S/genética , Recombinases , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Ovinos , Doenças dos Ovinos/diagnóstico , Fatores de Tempo
10.
MBio ; 10(2)2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-31015324

RESUMO

Murine studies suggest that the presence of some species of intestinal helminths is associated with changes in host microbiota composition and diversity. However, studies in humans have produced varied conclusions, and the impact appears to vary widely depending on the helminth species present. To demonstrate how molecular approaches to the human gut microbiome can provide insights into the complex interplay among disparate organisms, DNA was extracted from cryopreserved stools collected from residents of 5 rural Kenyan villages prior to and 3 weeks and 3 months following albendazole (ALB) therapy. Samples were analyzed by quantitative PCR (qPCR) for the presence of 8 species of intestinal parasites and by MiSeq 16S rRNA gene sequencing. Based on pretreatment results, the presence of neither Ascaris lumbricoides nor Necator americanus infection significantly altered the overall diversity of the microbiota in comparison with age-matched controls. Following ALB therapy and clearance of soil-transmitted helminths (STH), there were significant increases in the proportion of the microbiota made up by Clostridiales (P = 0.0002; average fold change, 0.57) and reductions in the proportion made up by Enterobacteriales (P = 0.0004; average fold change, -0.58). There was a significant posttreatment decrease in Chao1 richness, even among individuals who were uninfected pretreatment, suggesting that antimicrobial effects must be considered in any posttreatment setting. Nevertheless, the helminth-associated changes in Clostridiales and Enterobacteriales suggest that clearance of STH, and of N. americanus in particular, alters the gut microbiota.IMPORTANCE The gut microbiome is an important factor in human health. It is affected by what we eat, what medicines we take, and what infections we acquire. In turn, it affects the way we absorb nutrients and whether we have excessive intestinal inflammation. Intestinal worms may have an important impact on the composition of the gut microbiome. Without a complete understanding of the impact of mass deworming programs on the microbiome, it is impossible to accurately calculate the cost-effectiveness of such public health interventions and to guard against any possible deleterious side effects. Our research examines this question in a "real-world" setting, using a longitudinal cohort, in which individuals with and without worm infections are treated with deworming medication and followed up at both three weeks and three months posttreatment. We quantify the impact of roundworms and hookworms on gut microbial composition, suggesting that the impact is small, but that treatment of hookworm infection results in significant changes. This work points to the need for follow-up studies to further examine the impact of hookworm on the gut microbiota and determine the health consequences of the observed changes.


Assuntos
Anti-Helmínticos/administração & dosagem , Ascaríase/tratamento farmacológico , Bactérias/classificação , Microbioma Gastrointestinal/efeitos dos fármacos , Microbiota/efeitos dos fármacos , Necatoríase/tratamento farmacológico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Bactérias/genética , Criança , Pré-Escolar , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Fezes/microbiologia , Humanos , Quênia , Metagenômica , Pessoa de Meia-Idade , RNA Ribossômico 16S/genética , Reação em Cadeia da Polimerase em Tempo Real , População Rural , Análise de Sequência de DNA , Adulto Jovem
11.
PLoS Genet ; 15(4): e1008026, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30933976

RESUMO

Phosphorothioate (PT) modifications of the DNA backbone, widespread in prokaryotes, are first identified in bacterial enteropathogens Escherichia coli B7A more than a decade ago. However, methods for high resolution mapping of PT modification level are still lacking. Here, we developed the PT-IC-seq technique, based on iodine-induced selective cleavage at PT sites and high-throughput next generation sequencing, as a mean to quantitatively characterizing the genomic landscape of PT modifications. Using PT-IC-seq we foud that most PT sites are partially modified at a lower PT frequency (< 5%) in E. coli B7A and Salmonella enterica serovar Cerro 87, and both show a heterogeneity pattern of PT modification similar to those of the typical methylation modification. Combining the iodine-induced cleavage and absolute quantification by droplet digital PCR, we developed the PT-IC-ddPCR technique to further measure the PT modification level. Consistent with the PT-IC-seq measurements, PT-IC-ddPCR analysis confirmed the lower PT frequency in E. coli B7A. Our study has demonstrated the heterogeneity of PT modification in the bacterial population and we also established general tools for rigorous mapping and characterization of PT modification events at whole genome level. We describe to our knowledge the first genome-wide quantitative characterization of PT landscape and provides appropriate strategies for further functional studies of PT modification.


Assuntos
DNA Bacteriano/química , DNA Bacteriano/metabolismo , Fosfatos/metabolismo , Sequência de Bases , Sítios de Ligação/genética , DNA Bacteriano/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Genoma Bacteriano , Sequenciamento de Nucleotídeos em Larga Escala , Iodo , Reação em Cadeia da Polimerase , Salmonella enterica/genética , Salmonella enterica/metabolismo , Análise de Sequência de DNA , Espectrometria de Massas em Tandem
12.
Ecotoxicol Environ Saf ; 177: 93-99, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-30974248

RESUMO

In order to better understand the composition and driving factors of the bacterial community in Mollisols, we selected 9 representative facility agricultural lands in Mollisol area of China for sampling, and described it on a larger spatial scale. Soil bacterial community structure in these 9 regions (determined by high-throughput sequencing analysis) showed significant differences at the genus level. The correlation between bacterial community composition and soil properties, contaminants and geographical latitude showed that the diversity of bacterial community was still strongly correlated with pH and SOM under the influence of phthalates (P < 0.05). Principal component Analysis (PCA) showed that soil properties (i.e. pH, organic matter, stacking density, the content of nitrogen, potassium, phosphorus) and PAEs level rather than geographic latitude were main drivers of differences in bacterial community structure. These factors account for 73.04% of the total variation of the bacterial community. Among them, PAEs act as a typical pollutant is the main factor driving the composition of bacterial community in facility agriculture Mollisols. This shows that PAEs is a potential pollution risk factor, which has important guiding significance for the sustainable and healthy development of agriculture in Mollisol area.


Assuntos
Poluição Ambiental/análise , Ácidos Ftálicos/análise , Microbiologia do Solo , Poluentes do Solo/análise , Agricultura , Bactérias/classificação , Bactérias/efeitos dos fármacos , Bactérias/genética , China , DNA Bacteriano/química , Microbiota , Nitrogênio/análise , Fósforo/análise , Ácidos Ftálicos/toxicidade , Análise de Componente Principal , Análise de Sequência de DNA , Solo/química , Poluentes do Solo/toxicidade
13.
J Chem Phys ; 150(14): 144908, 2019 Apr 14.
Artigo em Inglês | MEDLINE | ID: mdl-30981230

RESUMO

We showed in our previous studies that just 3% cross-links (CLs), at special points along the contour of the bacterial DNA, help the DNA-polymer to get organized at micron length scales [T. Agarwal et al., J. Phys.: Condens. Matter 30, 034003 (2018) and T. Agarwal et al., EPL (Europhys. Lett.) 121, 18004 (2018)]. In this work, we investigate how does the release of topological constraints help in the "organization" of the DNA-polymer. Furthermore, we show that the chain compaction induced by the crowded environment in the bacterial cytoplasm contributes to the organization of the DNA-polymer. We model the DNA chain as a flexible bead-spring ring polymer, where each bead represents 1000 base pairs. The specific positions of the CLs have been taken from the experimental contact maps of the bacteria Caulobacter crescentus and Escherichia coli. We introduce different extents of ease of release of topological constraints in our model by systematically changing the diameter of the monomer bead. It varies from the value where chain crossing can occur freely to the value where chain crossing is disallowed. We also study the role of compaction of the chain due to molecular crowders by introducing an "effective" weak Lennard-Jones attraction between the monomers. Using Monte Carlo simulations, we show that the release of topological constraints and the crowding environment play a crucial role to obtain a unique organization of the polymer.


Assuntos
Cromossomos Bacterianos , Biopolímeros/química , Caulobacter crescentus/genética , DNA Bacteriano/química , Escherichia coli/genética , Modelos Biológicos , Método de Monte Carlo
14.
BMC Bioinformatics ; 20(1): 213, 2019 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-31029080

RESUMO

BACKGROUND: Next Generation Sequencing (NGS) is a commonly used technology for studying the genetic basis of biological processes and it underpins the aspirations of precision medicine. However, there are significant challenges when dealing with NGS data. Firstly, a huge number of bioinformatics tools for a wide range of uses exist, therefore it is challenging to design an analysis pipeline. Secondly, NGS analysis is computationally intensive, requiring expensive infrastructure, and many medical and research centres do not have adequate high performance computing facilities and cloud computing is not always an option due to privacy and ownership issues. Finally, the interpretation of the results is not trivial and most available pipelines lack the utilities to favour this crucial step. RESULTS: We have therefore developed a fast and efficient bioinformatics pipeline that allows for the analysis of DNA sequencing data, while requiring little computational effort and memory usage. DNAscan can analyse a whole exome sequencing sample in 1 h and a 40x whole genome sequencing sample in 13 h, on a midrange computer. The pipeline can look for single nucleotide variants, small indels, structural variants, repeat expansions and viral genetic material (or any other organism). Its results are annotated using a customisable variety of databases and are available for an on-the-fly visualisation with a local deployment of the gene.iobio platform. DNAscan is implemented in Python. Its code and documentation are available on GitHub: https://github.com/KHP-Informatics/DNAscan . Instructions for an easy and fast deployment with Docker and Singularity are also provided on GitHub. CONCLUSIONS: DNAscan is an extremely fast and computationally efficient pipeline for analysis, visualization and interpretation of NGS data. It is designed to provide a powerful and easy-to-use tool for applications in biomedical research and diagnostic medicine, at minimal computational cost. Its comprehensive approach will maximise the potential audience of users, bringing such analyses within the reach of non-specialist laboratories, and those from centres with limited funding available.


Assuntos
Biologia Computacional/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Interface Usuário-Computador , Esclerose Amiotrófica Lateral/genética , Esclerose Amiotrófica Lateral/patologia , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , Bases de Dados Factuais , HIV-1/genética , Humanos , Mutação INDEL , Polimorfismo de Nucleotídeo Único , RNA Viral/química , RNA Viral/genética , RNA Viral/metabolismo , Sequenciamento Completo do Genoma
15.
Curr Microbiol ; 76(6): 659-665, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30937514

RESUMO

In the present study, three strains (ChDC F213T, ChDC F251, and ChDC F267) were classified as novel species of genus Fusobacterium based on average nucleotide identity (ANI) and genome-to-genome distance (GGD) analysis and chemotaxonomic characterization. 16S rDNA sequences of strains ChDC F213T, ChDC F251, and ChDC F267 were highly similar to that of F. periodonticum ATCC 33693T (99.6, 99.4, and 99.4%, respectively). ANI and GGD values of the three isolates with F. periodonticum ATCC 33693T ranged from 92.5 to 92.6% and 47.7 to 48.2%, respectively. Considering that threshold of ANI and GGD values for bacterial species discrimination are 95-96% and 70%, respectively, these results indicate that the three isolates represent a novel Fusobacterium species. DNA G + C contents of the three isolates were 28.0 mol% each. Cellular fatty acid analysis of these strains revealed that C14:0, C16:0, and C16:1 ω6c/C16:1 ω7c were major fatty acids. Therefore, these three strains are novel species belonging to genus Fusobacterium. Strain ChDC F213T (= KCOM 1259T = KCTC 5677T = JCM 33009T) is the type strain of a novel species of genus Fusobacterium, for which a name of Fusobacterium pseudoperiodonticum sp. nov. is proposed.


Assuntos
Fusobacterium/classificação , Fusobacterium/isolamento & purificação , Boca/microbiologia , Composição de Bases , Análise por Conglomerados , Citosol/química , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Ácidos Graxos/análise , Fusobacterium/química , Fusobacterium/genética , Humanos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Homologia de Sequência do Ácido Nucleico
16.
Appl Microbiol Biotechnol ; 103(11): 4525-4538, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30993384

RESUMO

Rhamnose is a high-value carbohydrate used in flavorings, aromatics, and pharmaceuticals. Current demand for rhamnose is filled through plant-based sources; however, microbially originated rhamnolipids have been proposed as an alternative source. A mixed microbial biofilm, cultured from a wastewater sludge, was found to comprise > 8 dry weight% rhamnose when provided volatile fatty acids as carbon source, and 24 dry weight% when given glucose. The latter rhamnose concentration is a fourfold higher production mass than the current plant-based origin and is competitive with yields from pure microbial cultures. The biofilm was characterized based on total carbohydrate production at varying nutrient levels, individual carbohydrate monomer production from varying organic acid substrates, and microbial community composition-based on 16s rRNA. Biofilm carbohydrate production was maximized at a C:N ratio of 28 (mol:mol). The production of rhamnose varied significantly based on carbon substrate; glucose had the greatest yield of rhamnose, followed by propionic acid, lactic acid, acetic acid, valeric acid, and butyric acid. Microbial community analysis indicated an abundance of organisms within the Xanthobacter genus, which is known to produce rhamnose as zeaxanthin rhamnoside. Rhamnose production was heavily correlated with ribose production (R2 = 0.96). Results suggest that mixed microbial biofilms could be a competitive source of monomeric rhamnose that may be produced from mixed organic waste streams of variable composition via volatile fatty acids and glucose.


Assuntos
Biofilmes/crescimento & desenvolvimento , Consórcios Microbianos , Ramnose/metabolismo , Xanthobacter/crescimento & desenvolvimento , Xanthobacter/metabolismo , Carbono/metabolismo , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Ácidos Graxos/metabolismo , Glucose/metabolismo , Filogenia , RNA Ribossômico 16S/genética , Ramnose/isolamento & purificação , Análise de Sequência de DNA , Esgotos/microbiologia
18.
Curr Microbiol ; 76(6): 713-722, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30968206

RESUMO

Despite the broad assessment of sponge bacterial diversity through cultivation-independent and dependent strategies, the knowledge focusing on cultivable anaerobes from this holobiont is still incipient. Plakina is a genus with the highest number of described species from the smallest of poriferan classes, Homoscleromorpha. The Brazilian Atlantic coast has been presenting itself as a hotspot for the discovery of new plakinidae species, with initial surveys just now concerning to characterize their microbiome. The current study aimed to isolate and identify strict anaerobes from recently described species of Plakina collected at the coast of Cabo Frio, RJ. Samples of four sympatric morphotypes of Plakina cyanorosea and Plakina cabofriense were collected on the coast of Cabo Frio, RJ. Using five different culture media, a total of 93 bacterial isolates were recovered, among which 60 were strict anaerobes and, ultimately, 34 remaining viable. A total of 76.5% from these strains were mostly identified as Clostridium bifermentans by mass spectrometry and 82.4% identified by 16S rRNA sequencing, almost all of them affiliated to the genus Paraclostridium, and with one isolate identified as Clostridium butyricum by both techniques. None of the anaerobic bacteria exhibited antimicrobial activity by the adopted screening test. The present work highlights not only the need for cultivation and characterization of the anaerobic microbiota from marine sponges but also adds the existing scarce knowledge of culturable bacterial communities from Homoscleromorph sponges from Brazilian coast.


Assuntos
Bactérias Anaeróbias/classificação , Bactérias Anaeróbias/isolamento & purificação , Clostridiales/classificação , Clostridiales/isolamento & purificação , Poríferos/microbiologia , Aerobiose , Anaerobiose , Animais , Anti-Infecciosos/metabolismo , Organismos Aquáticos/microbiologia , Oceano Atlântico , Bactérias Anaeróbias/química , Bactérias Anaeróbias/genética , Técnicas Bacteriológicas , Brasil , Clostridiales/química , Clostridiales/genética , Clostridium bifermentans , Clostridium butyricum , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Espectrometria de Massas , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
19.
Nat Commun ; 10(1): 1153, 2019 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-30858373

RESUMO

Bacterial RNA polymerase employs extra-cytoplasmic function (ECF) σ factors to regulate context-specific gene expression programs. Despite being the most abundant and divergent σ factor class, the structural basis of ECF σ factor-mediated transcription initiation remains unknown. Here, we determine a crystal structure of Mycobacterium tuberculosis (Mtb) RNAP holoenzyme comprising an RNAP core enzyme and the ECF σ factor σH (σH-RNAP) at 2.7 Å, and solve another crystal structure of a transcription initiation complex of Mtb σH-RNAP (σH-RPo) comprising promoter DNA and an RNA primer at 2.8 Å. The two structures together reveal the interactions between σH and RNAP that are essential for σH-RNAP holoenzyme assembly as well as the interactions between σH-RNAP and promoter DNA responsible for stringent promoter recognition and for promoter unwinding. Our study establishes that ECF σ factors and primary σ factors employ distinct mechanisms for promoter recognition and for promoter unwinding.


Assuntos
Proteínas de Bactérias/química , RNA Polimerases Dirigidas por DNA/química , Regulação Bacteriana da Expressão Gênica , Mycobacterium tuberculosis/genética , Fator sigma/química , Iniciação da Transcrição Genética , Proteínas de Bactérias/metabolismo , Cristalografia por Raios X , DNA Bacteriano/química , DNA Bacteriano/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Holoenzimas/química , Holoenzimas/metabolismo , Modelos Moleculares , Regiões Promotoras Genéticas/genética , Fator sigma/metabolismo
20.
Nat Commun ; 10(1): 972, 2019 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-30814507

RESUMO

Bacterial toxin-antitoxin (TA) modules are tightly regulated to maintain growth in favorable conditions or growth arrest during stress. A typical regulatory strategy involves the antitoxin binding and repressing its own promoter while the toxin often acts as a co-repressor. Here we show that Pseudomonas putida graTA-encoded antitoxin GraA and toxin GraT differ from other TA proteins in the sense that not the antitoxin but the toxin possesses a flexible region. GraA auto-represses the graTA promoter: two GraA dimers bind cooperatively at opposite sides of the operator sequence. Contrary to other TA modules, GraT is a de-repressor of the graTA promoter as its N-terminal disordered segment prevents the binding of the GraT2A2 complex to the operator. Removal of this region restores operator binding and abrogates Gr aT toxicity. GraTA represents a TA module where a flexible region in the toxin rather than in the antitoxin controls operon expression and toxin activity.


Assuntos
Toxinas Bacterianas/genética , Toxinas Bacterianas/metabolismo , Antitoxinas/genética , Antitoxinas/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/toxicidade , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , Genes Bacterianos , Proteínas Intrinsicamente Desordenadas/química , Proteínas Intrinsicamente Desordenadas/genética , Proteínas Intrinsicamente Desordenadas/metabolismo , Modelos Moleculares , Conformação de Ácido Nucleico , Óperon , Regiões Promotoras Genéticas , Ligação Proteica , Dobramento de Proteína , Estrutura Quaternária de Proteína , Pseudomonas putida/genética , Pseudomonas putida/metabolismo , Eletricidade Estática , Sistemas Toxina-Antitoxina/genética
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